1. Field of the Invention
The invention is on the compositions and uses of the extract from the annatto seed and such extract that is annatto oil or oleoresin containing non-saponifiables, especially non-saponifiable terpenoids.
2. Description of the Related Art
Tocotrienols generally are classified as farsnesylated chromanols (FC) and mixed terpenoids (US 2004-0202740 A1, Tan). Tocopherol and tocotrienol are believed to have beneficial effects because they act as antioxidants. Tocotrienols, in particular, have been documented to possess hypocholesterolemic effects as well as an ability to reduce atherogenic apolipoprotein B and lipoprotein plasma levels. Further, tocotrienols are believed to be useful in the treatment of cardiovascular disease and cancer (Theriault, A., et al., “Tocotrienol: A Review of its Therapeutic Potential,” Clinical Biochemistry, 32:309-319 (July 1999); and “Tocotrienols: Biological and Health Effects,” in Antioxidant Status, Diet, Nutrition, and Health, Papas, ed. (CRC Press), pp. 479-496 (1999)). Delta-tocotrienol and gamma-tocotrienol, in particular, have been identified as effective suppressants of cholesterol activity (Qureshi, et al., “Response of Hypercholesterolemic Subjects to Administration of Tocotrienols,” Lipids, 30(12) (1995)), and in inducing apoptosis of breast cancer cells (Yu, et al., “Induction of Apoptosis in Human Breast Cancer Cells by Tocopherols and Tocotrienols,” Nutrition and Cancer, 33(1):26-32 (1999)).
Tocols, which includes tocopherols and tocotrienols, have several sources, including several vegetable oils, such as rice bran, soybean, sesame and palm oils. Tocotrienols have been discovered in the seeds of Bixa orellana Linn, otherwise known as the achiote tree (Jondiko, I. S., et al., “Terpenoids and an Apocarotenoid from Seeds of Bixa Orellana,” Phytochemistry, 28(11):3159-3162 (1989)). However, each source of tocotrienols and tocopherols generally contains more than a single tocol homolog. For example, palm oil and rice bran oil generally include both tocotrienols and tocopherols. Further, alpha-tocopherol has been reported to attenuate certain effects of tocotrienols, such as the cholesterol-suppressive activity of gamma-tocotrienol (Qureshi, et al., supra.). In addition, because of their structural similarity, tocotrienols and tocopherols can be difficult to separate.
Geranyl geraniol (GG) includes acyclic diterpene alcohols and geranyl geraniated terpenoids, and occurs naturally in linseed oil and cedrela toona wood and tomato fruit. Geranyl geraniol also has been discovered to exist in the seeds of Bixa orellana (Craveiro, et al., “The Presence of Geranyl geraniol in Bixa Orellana Linn,” Quimica Nova, 12(3):297-298 (1989)). Potential uses for geranyl geraniol include synthesis of co-enzyme Q10, vitamin K and tocotrienols. It is believed to inhibit esterification of retinol into inactive retinyl esters and, therefore, may be used to improve skin desquamation and epidermal differentiation (U.S. Pat. No. 5,756,109, issued to Burger, et al. on May 26, 1998). Geranyl geraniol has been employed in conjunction with HMG-CoA reductase inhibitors in treatment of elevated blood cholesterol (WO 99/66929 by Scolnick, published Dec. 29, 1999). Geranyl geraniol also is suspected to be useful for treatment of human prostate cancer (U.S. Pat. No. 5,602,184, issued to Myers, et al. on Feb. 11, 1997).
Bixa orellana Linn, otherwise known as the achiote tree, is a member of the Bixaceae family and is native to tropical America. It is grown commercially in other parts of the world, generally within 20° of the equator or more preferably within 15° of the equator. The seeds of Bixa orellana Linn are the source of a reddish-orange colorant, known as annatto, that contains bixin and orelline, both of which are carotenoid pigments. The colorant is used commonly in foods, dyes and polishes. Typically, annatto is extracted from dehusked seeds in an aqueous caustic solution. The colorant is precipitated from aqueous caustic solution by addition of a suitable acid, such as sulfuric acid. The precipitated colorant is removed by filtration. Filtercake of precipitated annatto colorant is dried and milled to form a commercial product. An oily phase generally is separated from an aqueous caustic phase by centrifugation or by settling. Alternatively, the annatto colorant can be extracted from seeds in an organic solvent, such as hexane, acetone, or an alcohol. Miscella containing color and byproduct oil are allowed to cool sufficiently to precipitate the annatto colorant. The precipitate is separated as bottoms from the organic solvent. The oily phase from the caustic or organic extractions following separation of the annatto precipitate generally are discarded as byproducts.
The phrase “annatto extract” is imprecise and is used by other authors to define a wide array of compositions. Typically, these products contain all the parts of the annatto seed or at least contain the bixins that are used for colorants. It is important to distinguish annatto extracts that contain bixins from the disclosed annatto extracts by the applicants that do not contain bixins or are essentially free of bixins.
U.S. Pat. No. 6,350,453 by Tan discloses the method of obtaining the natural annatto extract, the byproduct solution of Bixa orellana seed components. The final product of the method is a composition containing the natural ratio of isomers of the principal components.
The disclosed method in Tan('453) uses a “a vegetable oil” or “a rice bran oil” (Column 4, Lines 34-42) in one example to “reduce viscosity” (Line 39). This vegetable oil or rice bran oil was a cooking oil (triglyceride (TG)) which contained traces or none of any tocotrienols, tocopherols or geranyl geraniols.
The vegetable oil was used as a lubricant for the distillation of annatto extract as it becomes more viscous with successive distillations (or “passes”). This is shown in Examples 3 & 4 in Tan('453) with the use of 10% rice bran oil.
Careful Examination of Example 4 in Tan('453), going over Passes 1 and 2, after rice bran oil was added, then the Pass 2 distillation, tocotrienol was enriched. The vegetable oil was never distilled with the tocotrienol because the rice bran oil served as a lubricant (viscosity reducer) and not an additive or diluent to the tocotrienol. The tocotrienol enrichment (about 15% to 26.3% in Pass 2, Example 4 or about 19.6% to 33.6% in Pass 2, Example 3) shows that the vegetable oil was left behind as residue and did not distill with tocotrienol. The difference in molecular weights of TG and tocotrienol made this processing possible. In Example 3, the rice bran oil ended in residue of Pass 2 (accounting 32% of the total mass).
The byproduct solution of Bixa orellana seed components is obtained after removing the bixins to produce “yellow cake” and has greatly reduced levels of bixins. The byproduct solution is then distilled to obtain a 290-390 Dalton MW fraction. The 290-390 Dalton MW fraction is the fraction that contains geranyl geraniol. The disclosed geranyl geraniols in the application are obtained from the 290-390 Dalton MW fraction of the byproduct solution of Bixa orellana seed components.
Geranyl geraniols are 290 Daltons in molecular weight and Bixins are 390-425 Daltons in molecular weight. These molecular weights are easy to calculate from the structures of these chemicals. Additionally, their chemical structures show that geranyl geraniols have 1 oxygen groups and bixins have 4 oxygen groups.
Although the molecular weights of bixins (390-425 Daltons) over lap the molecular weights of tocotrienol and tocopherol (350-450 Daltons), their chemical structure [bixins have 4 oxygen groups; tocotrienols and tocopherols have 2 oxygen groups] inhibits bixins (i.e., they are heavier because they contain the 4 large oxygen groups) distillation from the byproduct solution of Bixa orellana seed components and remain in the residue material The higher the number of oxygen groups a molecule contains, the less likely it is to distill. Geranyl geraniol has 1 oxygen group, tocotrienols and tocopherols have 2 oxygen groups, bixins have 4 oxygen groups; and triglycerides have 6 oxygen groups (MW 500-1,000 Daltons). Therefore, geranyl geraniol distills before the tocotrienols and tocopherols, and bixins and triglycerides (being the heaviest/densest) remain in the residue.
It has been discovered that byproduct solutions of Bixa orellana seed components contain tocotrienols, including delta- and gamma-tocotrienols, and geranyl geraniol. In particular, it has been discovered that tocotrienols and geranyl geraniol are present in the byproduct oily phase of annatto colorant from annatto seeds and, especially, from whole dehusked annatto seeds.
A “byproduct solution of Bixa orellana seed components” is defined herein as a solution derived from Bixa orellana seed components having a concentration of annatto colorant significantly reduced from that of Bixa orellana seeds themselves. Other common terms for byproduct solution used for commercial products include: oil-soluble annatto color or annatto oil. Generally, the concentration of annatto colorant, which is defined as bixins and other carotenoids, chemically modified, altered or esterified, in byproduct solution of Bixa orellana seed is less than about two percent, by weight, such as between about 0.05 weight percent and about 2.0 weight percent.
Annatto extract composition (AEC) typically contains cis and trans isomers of geranyl geraniol (GG) and tocopherol-free tocotrienols (T3) that are essentially delta and gamma isomer forms. Geranyl geraniols belong to a class of terpenoid, more specifically, diterpene isoprenoids containing four isoprene units. The GG may be all in the trans isomer form (only one form possible), and/or contain one or more of cis isomer forms, both of the trans and cis forms are endogenous nutrients; however, they are not vitamins in the classical sense. Both cis and trans GG become substrates for many branch-point reactions needed in the syntheses of downstream isoprenoid and distal protein products.
Geranyl geraniol (cis and trans) has a molecular weight of 290 Daltons, which is much smaller than the tocopherols and tocotrienols, and bixins in the annatto seed. Vitamin E, including tocopherols and tocotrienols, are typically 390-430 Daltons in molecular weight or more broadly 350-450 Daltons in molecular weight, which includes tocopherols and tocotrienols without any methylated groups in the lower range and tocopherols and tocotrienols with fully methylated groups in the higher range. [Alpha-Tocopherol=430, Beta-Tocopherol=417, Gamma-Tocopherol=417, Delta-Tocopherol=403, Alpha-Tocotrienol=424, Beta-Tocotrienol=411, Gamma-Tocotrienol=410, and Delta-Tocotrienol=396.]
Many physiologic nutrients of small molecular weight are produced from the mevalonate pathway that generates the “isoprenoid pool” (IP) products. Geraniol (G), farnesol (F), and GG are the examples of IP products containing two, three, and four repeating units of five-carbon isoprenes, respectively. Tocotrienols belong to the class of vitamin E that includes tocopherols. It is known that T3's lower cholesterol and treat hypercholesterolemia (Pearce, Parker et al. 1992; Song, DeBose-Boyd 2006). Unlike GG, T3's are not endogenous nutrients, but are produced by plants and have a condensed farnesol tail in its structure.
Farnesol constitutes the last committed step to cholesterol synthesis, but GG is not required for cholesterol synthesis (Flint, Masters et al. 1997; Flint, Masters et al. 1997). GG constitutes the first uncommitted step to cholesterol synthesis, and therefore, the first committed steps in the synthesis of CoQ10, dolichol (DL), heme porphyrin, and GG-prenylated and DL-glycosylated proteins (Baker and Tarnopolsky 2001). Both cis and trans isomeric GGs are required for endogenous isoprenoid substrates for downstream branch-point products (Grünler, Ericsson et al. 1994). Trans-GG is the precursor to all-trans CoQ10 synthesis, which is involved in mitochondrial respiration. Cis-GG is the precursor to DL, DL-glycosylated proteins, and certain GG-prenylated proteins. Dolichol and GG tend to concentrate in the brain and liver but GG is ubiquitously found in many tissues (Grünler, Ericsson et al. 1994). Proteins produced by DL-glycosylation and GG-prenylation will be directed (e.g., structures of protein fold, targets of where it will be delivered, and anchors of how it will be recognized). Deficiency in GG and/or DL leads to improper localization of proteins, producing nonsense proteins and signals. A major use of GG-prenylated protein is in the muscle tissues, and a major use of DL-glycosylated protein is in the nerve tissues. Synthesized proteins via isoprenoid GG and DL are described.
The HMG CoA reductase (HMGR) catalyzes the rate-limiting steps in the lengthy hepatic cholesterol synthesis. The inhibition of HMGR is the target for statin targetment of hypercholesterolemia. However, statins inhibit mevalonate (e.g., one isoprene) at the onset of the formation of the first isoprene, and therefore inhibits all subsequent IP products, including GG (FIG. 1). It is this depletion and deprivation of GG that can produce secondary, but clinically significant, side effects of DL-starved cranial nerve damage and defects typified by neurological dysfunctions (e.g., taste alteration/loss, lack coordination, facial paresis, memory loss, vertigo, peripheral neuropathy, and peripheral nerve palsy). Geranyl geraniol salvages GG-prenylated proteins in brain cells (That, S. Rush et al. 1999). Brain cells utilize free GG (not in the activated GG-diphosphate form: GGPP) to restore the IP pool and incorporate it into the protein biosynthesis system. Thus, GG is physiologically and pharmacologically significant in the central nervous system (CNS) (Kotti, T. J., D. M. Ramirez, et al 2006; Sever, N. B. Song, et al 2003; Bi, X., M. Baudry, et al 2004). For example, when isoprenoid products are depleted by statin and bisphosphonate medication, GG replenishes GG-prenylated and DL-glycosylated proteins. Drug side effects are many and they include GG-deprived induction of myotoxicities (e.g., musculoskeletal disorders, muscle cramps/pain, myalgia, myopathy, rhabdomyolysis, and myonecrosis), exo- and endothelial dysfunctions (e.g., upper GI maladies—esophagitis, gastritis/stomatitis, stomach/duodenal ulcer and lower GI maladies—constipation, dyspepsia, gastric dysmotility, abdominal pain) (Watts, Freedholm et al. 1999). GI tract (i.e., esophageal, gastric, duodenal) lesions include perforations, ulcers, bleeds and hemorrhages, maladies all of which come from GG-deprived protein synthesis of the mucosae. Other GG-deprived dysfunctions include ocular maladies (e.g., cataract/lens opacity, dry eyes, corneal abrasion, ophthalmoplegia), anemia, CoQ10, DL and its associated DL-starved maladies, described above. Again, eye problems such as lens opacity and dry eyes can be traced to the deprivation of GG. These side effects include secondary CoQ10-deprived maladies (e.g., mitochondrial dysfunction, ATP/respiration, LDL protection, tiredness/malaise, fibromyalgia, chronic fatigue syndrome, and congestive heart failure). The schematic outline of this invention for GG-deprived maladies is shown in FIG. 2.
Drug-Induced Myopathies Via GG Inhibition
IP product depletion from treatment with statins is serious side effect, so alternatives to statins are proposed for treatment of hypercholesterolemia. Squalene synthase catalyses the first committed step in cholesterol biosynthesis via two F groups head-to-head (FIG. 1). To avoid such global IP depletion, and particularly GG depletion, squalene synthase inhibitors (SSI) target distal isoprenoid squalene inhibition to treat hypercholesterolemia (Ciosek, Magnin et al. 1993; Amin, Rutledge et al. 1997). A unique advantage of SSI, as opposed to statins, is that they do not deplete IP immediate and distal products, such as GG, CoQ10, and DL. Such new drug targets only underscore the unique role of GG and the serious implication of its depletion. However, widespread successful use of statins, and their ever growing expanded uses, emphasizes the importance of the invention for adjunctive therapy to circumvent isoprenoid depletion in general, and GG depletion in particular (Johnson, T. E., X. Zhang, et al 2004).
Isoprenoid pool deprivation and myopathies are common with widespread use of statin drugs for the treatment of hypercholesterolemia, fibrate drugs for the treatment of hypertriglyceridemia, and bisphosphonate drugs for the treatment of osteoporosis. Such widespread use of statins is now extended further because of other non-cholesterol approved uses, other cardiovascular indications/uses, as well as, other statin-in-tandem combination uses. A clinically meaningful adverse event of GG inhibition is a global loss of protein, with consequent myotoxicity. Therefore, AEC is particularly useful in the adjunctive relief to IP deprivation, such as, but not limited to statin, fibrate, and bisphosphonate users.
Non-Drug-Induced Myopathies Via GG Inhibition
Isoprenoid pool deprivation may also occur in the elderly and those with AIDS-HIV where wasting occurs due to protein deficit (Poels and Gabreels 1993; Hamilton-Craig 2001).
CoQ10
CoQ10 is transported in the vascular system via LDL particles. Statins work to inhibit de novo cholesterol synthesis, which also simultaneously inhibit de novo CoQ10 and DL synthesis (Bliznakov 2002). Statins also work to increase the hepatic LDL receptors, hence reducing LDL particles in vascular circulation. Consequently, patients on statins will see a drop in LDL with a corresponding drop in CoQ10 (Watts, Castelluccio et al. 1993).
GG is the first committed step for numerous downstream distal products, including CoQ10 (FIGS. 1 & 4). The GG molecule (MW=290) containing 4 isoprene units anabolizes to CoQ10 molecule (MW=863) containing 10 isoprene units. Conceptually, a minimum of 2 moles of GG is required to anabolize 1 mole of CoQ10 and conversely 1 mole of CoQ10 is required to catabolize to 2 moles of GG. This is illustrated by way of the molar conversion example as follows: A 100 mg of GG (100/290=0.345 mmole) can anabolize to 150 mg of CoQ10 (0.345/2×863).
Hypercholesterolemia
Statin intensifies in vivo LDL oxidation in patients with myocardial ischemia while CoQ10 supplementation suppresses lipid oxidation (Lankin, Tikhaze et al. 2000). Further, animal cells contain about 10-fold more CoQ10 than vitamin E, and the cell preferentially utilizes CoQ10 as an antioxidant.
This invention shows AEC supplementation prevents statin toxicities, increases CoQ10, and the endogenous CoQ10 preferentially protects the LDL, lowers cholesterol and improves endothelial functions all at the same time. For patients on statins, endogenous CoQ10 levels typically drop about 30-40%. Clinically significant adverse effects occur when CoQ10 levels fall below 0.5 ug/mL. AEC also help diabetics on statins by enhancing CoQ10 status which improves beta-cell function in Type 2 diabetes (McCarty 1999).
It is implicit to current discussions that GG is readily bioavailable to cells and tissues. In addition, GG is not cytotoxic as it does not cause cell rounding, a known cellular indicator of myotoxicity (McGuire and Sebti 1997; Ownby and Hohl 2002). In fact, GG prevents and reverses cell rounding caused by statins and bisphosphonates. However, a similar IP product, farnesol, does not have either of these GG benefits. Therefore, the use of AEC takes advantage of the bioavailability and safety of GG to tissues.
Statin inhibits the insulin-responsive glucose transporter (Glut 4), and that such inhibition of IP biosynthesis cause IR in adipocytes (Chamberlain 2001). Glut 4 is a membrane protein that requires GG-prenylation. Therefore, the use of statins and bisphosphonates would inhibit the GG-prenylated biosynthesis of Glut 4, and thereby causing insulin resistance (IR) in adipocytes.
Cancer
A strategic way to inhibit cancer is to employ a farnesyl transferase inhibitor (FTI), since Ras cancer requires farnesyl-prenylation of its protein for survival. These FTIs are known to have toxic effects to cancer patient including GI toxicity, peripheral neuropathy & nerve conduction abnormality, and fatigue (Johnston and Kelland 2001). Surprisingly, all of these toxic effects may be ascribed to GG deficiency. GI toxicity is due in part to GG-associated prenylation of protein on the GI lining. Neuropathy and nerve defects are often related to DL-depleted glycosylation. Fatigue is often of unknown etiologies, commonly associated with chronic fatigue syndromes. They are ascribed to a deficiency in CoQ10, derived endogenously from the GG substrate.
Statin drugs have also been used in cancer treatment. A typical dosage of statins for cancer is 10 times their requirements for cholesterol reduction (Wong, Dimitroulakos et al. 2002). This can lead to serious myotoxicities including myopathy and rhabdomyolysis. GG is not toxic to untransformed cells or to normal cells (Stark, Blaskovich et al. 1998; Ownby and Hohl 2002; Wong, Dimitroulakos et al. 2002).
Cancer patients often have low blood levels of CoQ10. CoQ10 has been used as treatment in patients with breast and prostate cancers (Folkers, Osterborg et al. 1997; Judy, Nguyen et al. 2004). The prostate specific antigen (PSA) and prostate mass of prostate cancer patients after one year of CoQ10 supplementation decreased 71% and 47%, respectively. However, the mechanism of such effect is not yet known. Prostate cancer patients taking up to 600 mg/day CoQ10 is equivalent to taking 400 mg/day supplement of GG (Judy, Nguyen et al. 2004) according to earlier analysis (see CoQ10 section).
CoQ10 reduces the severity but not the incidence of musculoskeletal toxicities and patient complaints (Thibault, Samid et al. 1996; Wong, Dimitroulakos et al. 2002). Supplementation of mevalonate, a direct precursor to GG but not CoQ10, is shown to ameliorate myopathy, suggesting that the toxic effects are not due to CoQ10 deficiency (Smith, Eydelloth et al. 1991). These studies lend corroborative support to the above claim that CoQ10 catabolizes to GG, at least in parts, which in turn is responsible for partial reversal of myopathy. It may also be understood that it is GG, not CoQ10 per se, reverses myopathy.
While many biological processes are anabolic in nature, catabolic processes are also well known. One such isoprenoid catabolism is the conversion of cholesterol to Vitamin D, steroid hormones, and bile acids (FIG. 2). Such a strategy of cancer treatment is unique, as both CoQ10 and GG are endogenous nutrients, while the majority of cancer drugs are xenobiotic.
There are numerous strategies that disclose the use of GG for cancer treatment, which directly or indirectly involve GG protein prenylation (McGuire and Sebti 1997; Ownby and Hohl 2002). However, its apoptosis mechanism remains largely unknown. Two hypotheses come closest to explaining the mechanism as a “common effector” or a “coordinated regulator” of apoptosis by GG. GG results in a rapid en masse induction of apoptosis via activation of caspase-3 and possibly caspase-2 (Polverino and Patterson 1997). GG very quickly induces phosphatidyl choline biosynthesis inhibition at the level of choline phosphotransferase, the last step of CDP-choline known as the Kennedy pathway (Miquel, Pradines et al. 1998). Surprisingly, neither of the two apoptosis hypotheses require GG prenylation nor involve protein synthesis for apoptosis. GG appears to be the common denominator and a very potent compound to induce apoptosis en masse. It should be noted that GGPP is not stable and is unlikely to penetrate cell membranes unaided, but the natural isoprenol GG is bioavailable, and taken up by cells through an active transport system, and/or dephosphorylated sequentially by kinases (Danesi, McLellan et al. 1995; Bentinger, Grunler et al. 1998).
Renal Insufficiency
Renal insufficiency affects about 20 million Americans. The continuous irritation of the peritoneum in peritoneal dialysis patients can result in local peritoneal fibrinolytic activities as measured by fibrinolytic enzyme tissue-type plasminogen activator (t-PA) and plasminogen activator inhibitor-1 (PAI-1). Statins increase the t-PA and decrease the PAI-1 and may cause defects in the actin cytoskeleton (Haslinger, Goedde et al. 2002), which may irritate and thin the peritoneal lining. It is noted that the negative effects of statins can be prevented or reversed by the use of GG (Colli, Eligini et al. 1997; Haslinger, Goedde et al. 2002). Since many statins including cerivastatin, pravastatin, lovastatin, and simvastatin are filtered in part through the kidneys and excreted as urine, these drugs can exasperate the problems of renal insufficient patients.
Organ Transplants
Annually there are approximately 2,000 heart and 14,000 kidney transplants performed in the US. Patients with kidney and heart transplants are normally given cyclosporine to suppress the immune response to organ rejection. The most common side effects of cyclosporine are kidney dysfunction and failure, as measured by elevated blood creatinine and uric acid. These side effects may be caused by decreased efficiency in the glomerular filtration rate (GFR), indicating renal insufficiency. Since most graft patients have elevated lipid levels that can lead to coronary artery disease, statins are often prescribed along with cyclosporine. For these patients, the risks of myopathy and/or rhabdomyolysis are substantially higher (ca 15-80%). Despite the dangers of myotoxicities of this combo therapy, their usage is justified based on benefit-to-risk assessment provided that the statin doses are on the lower end, only one statin is allowed, and no fibrates (Ballantyne, Corsini et al. 2003).
Myotoxicities
Myotoxicity includes all forms and stages of muscle damage including, but not limited to, myalgia, myopathy, and rhabdomyolysis. Myopathy is also associated with generalized myalgia and recurrence of fatigue or weakness (creatine kinase level, CK >10 times the normal value). Rhabdomyolysis is characterized by global skeletal muscle fiber breakdown. Organ damage, typically renal insufficiency or acute renal failure, accompanies rhabdomyolysis when CK >100 times the normal value.
Myopathy and rhabdomyolysis may also have non-drug origins. Among the common causes that are not drug-induced are traumas (e.g. surgery), infections (e.g. viral, bacterial, and fungal), exercise exertion, alcohol abuse, and other inherited, environmental, or metabolic causes (Poels and Gabreels 1993; Hamilton-Craig 2001). Therefore, myotoxicity of both drug-induced and non-drug-induced origins are widespread as evidenced by the mild form, myalgia, to intermediate form, myopathy, to severest form, rhabdomyolysis.
There are many known causal mechanisms for drug-induced myopathies including inhibitions of cytochrome 3A4, HMGR, GG, and P-glycoproteins. Statins and bisphosphonates are particularly effective inhibitors of HMGR and GG. These two classes of drugs have remarkably overlapping modes of action. For example, statins, known for its cholesterol reduction via HMGR inhibition, reduce osteoporosis (Rogers 2000; Cruz and Gruber 2002). Conversely, bisphosphonates, known for bone strengthening via GG inhibition, reduce cholesterol (Ciosek, Magnin et al. 1993). Surprisingly, both statins and bisphosphonates inhibit cancer via FT inhibition (Luckman, Coxon et al. 1998; Wong, Dimitroulakos et al. 2002).
Most drugs are extensively biotransformed by the metallo-protein enzyme cytochrome P450 (CYP) system, with the majority of them processed by CYP 3A4, including statins. These processed drugs are removed from the body through biliary and renal excretions in a safe manner. When enzymatic processing by CYP 3A4 is depressed, drug concentration (e.g. statin) becomes elevated in the blood. Such elevation can occur during statin monotherapy or combo-therapy with erythromycin (where blood statin concentration is known to increase by 3-8 folds) (Ayanian, Fuchs et al. 1988; Spach, Bauwens et al. 1991) or with cyclosporine (where blood statin concentration is known to increase by 6-23 folds) (Regazzi, Iacona et al. 1993; Olbricht, Wanner et al. 1997; Holdaas, Jardine et al. 2001). Similar interactions can occur with other drug classes such as warfarin, antifungals/antibiotics, and niacin. The resultant statin elevation in the vascular system can cause serious GG depletion, leading to myopathy and rhabdomyolysis. It is important to note that GG does not inhibit any of the cytochrome P450 enzymes for which CYP3A4 is a part of (Raner, Muir et al. 2002).
Fibrates are effective in lowering triglyceride and hence are particularly useful for prediabetics and Type II diabetics; however, they tend to have a high toxic side effect of myopathy. For prediabetic and diabetic patients, benefits may outweigh the risk in combo therapy with statins to treat mixed lipidemia, common in this patient group. However, the incidence of myopathy may increase by 10-folds in diabetics as compared to the general population when on combo therapy (i.e. myopathy increased from 0.12% to 1.35%) (Gavish, Leibovitz et al. 2000; Omar, Wilson et al. 2001).
Even in monotherapy, fibrates cause myopathy 5.5 times greater than statins, posing an independent risk for myopathy. Fibrates are excreted through the kidneys, which can cause serious problems even in people with mild renal impairment.
Insulin Resistance
Insulin resistance (IR) is associated with increased risk of cardiovascular disease (CVD), Type 2 diabetes mellitus (T2DM), hypertension, polycystic ovarian syndrome (PCOS) and alcohol-unrelated fatty liver disease. However, plasma insulin measurement is not standardized across clinical laboratories, and therefore is an unreliable marker. Therefore, a surrogate marker was developed for insulin resistance, where the IR criteria are TG/HDL≧3.5 and/or TG≧140 mg/dL (McLaughlin, Abbasi et al. 2003).
GG activates mixed PPARs, both PPARγ at the adipocytes and PPARα at the hepatocytes (Takahashi, Kawada et al. 2002). PPARγ activation in adipose tissues decreases IR (Lehmann, Moore et al. 1995; Willson, Lambert et al. 2001) and PPARα activation in the liver lowers blood lipids (Peters, Hennuyer et al. 1997; Staels, Dallongeville et al. 1998). Furthermore, statin down regulates glucose transporter 4 (Glut 4) expression and thereby suppresses the glucose uptake into cells with consequent IR (Chamberlain 2001). Therefore, IP products that are decimated by statin inhibition may inhibit the GG-prenylated protein synthesis of Glut 4.
Peroxisomal Proliferator Activated Receptors
Peroxisomal proliferator activated receptors (PPAR) are members of the nuclear receptor transcription factors. The metabolic consequences of PPARγ activation have been researched mostly on adipose tissue where it is largely expressed (Kraegen 1998; Smith 1998), as well as, on muscle tissue (Hevener, He et al. 2003). The metabolic effects of known PPAR activator thiazolidinedones (TZD) are, a) reduces hyperglycemia and hyperinsulinemia, b) lowers FFA and TG levels, c) enhances IS and lowers IR states, and d) requires insulin for glucose-lowering action. Numerous PPARγ activator functions are similar to PPARα activator functions. This PPARα has been actively researched on liver tissue, especially with regards to lipid use (e.g., uptake and beta-oxidation). Even though the action sites of PPARγ (mainly in adipose) and PPARα (mainly in liver) are different, their activations have many overlapping outcomes. Typically TZD and fibrates affect the activation of PPARγ and PPARα, respectively.
Sterol Regulatory Element Binding Protein-1
Sterol regulatory element binding protein-1 (SREBP-1) is a transcription factor that responds to nutritional status and regulates metabolic gene expression in various organs, including liver, adipose and muscle. It has been shown that insulin and glucose induces de novo fatty acid synthesis leading to a rapid increase in lipogenic flux in skeletal muscle. This lipid accumulation is associated with muscle IR in obesity and T2DM, and is stimulated/mediated via the SREBP-1 expression (Guillet-Deniau 2003). As discussed earlier, IR is tightly associated with increased lipids (McLaughlin, Abbasi et al. 2003) and increased insulin or hyperinsulinemia (HI) (DeFronzo 1998). Additionally, the SREBP-1 expression in part controls FFA/TG synthesis, and PPAR expression in part controls FFA/TG uptake and catabolism (Song, B., R. A. DeBose-Boyd 2006).
Other Aspects of GG Deficiencies and Uses:
The upper GI track (esophagus, stomach, and duodenum) is particularly sensitive to perforations, ulcers, and bleeds. Collective adverse events (AE) include, but not limited to, abdominal pain, dyspepsia, esophageal erosion, esophagitis, reflux esophagitis, and the likes in the duodenum. Repairs to the GI track are done by cellular replication and take approximately 2 weeks in esophagus. Repairs by mucosal migration take approximately 2 days in the duodenum and 2 hours in the stomach. Therefore opportunistic AE is most likely to occur in the esophagus followed by duodenum and least likely in the stomach. Not surprisingly, drug-induced upper GI AE are common, especially in the esophagus. These drugs include emepronium bromide, doxycycline, tetracycline antibiotics, iron supplements, quinidine, non-steroid anti-inflammatory drugs (NSAIDs), alprenolol, captopril, theophylline, zidovudine, and bisphosphonates. Studies show 20-30% of patients develop upper GI AE within the first year of bisphosphonate therapy (Talley, Weaver et al. 1992). The mechanism of upper GI ulcer-related events is due to the GI's inability to prenylate protein needed for cellular replication (a much slower process than mucosal migration) caused by drug-induced depletion of GG and localized esophagitis caused by pills slipping through the esophagus (Watts, Freedholm et al. 1999)
Asymptomatic endoscopic abnormalities (e.g. hemorrhages, erosions, and ulcers) are surprisingly high (15%) in normal post menopausal women (Watts, Freedholm et al. 1999).
Steroids are widely used and the most common among them is prednisone. Corticosteroids are used for many inflammatory diseases including but not limited to arthritis, connective tissue disease, asthma, and in heart transplant patients. These corticosteroids have several side effects including rapid loss of bone mass in the first year of use, as high as 15% of patients develop vertebral fractures (Adachi and Ioannidis 2000), loss of bone mineral density even at very low doses, e.g. prednisone at 5 mg/day (Saito, Davis et al. 1995), and a high rate of steroid-induced osteoporosis, higher than osteoporosis in post menopausal women (Miller 2001). To prevent and reverse corticosteroid-induced osteoporosis, bisphosphonates has become the best drug candidate.
The role of Vitamin E in exercise is well known. Muscle damage can occur during exhaustive exercise, even in highly trained athletes. Furthermore, since the body's Vitamin E consumption increases with the amount of exercise, high amounts of Vitamin E are needed for endurance training and for membrane lipid oxidation protection during strenuous exercise (VERIS 1989).
Statins and bisphosphonates can increase the risk of adverse ocular side effects including cataracts (Schlienger, Haefeli et al. 2001). Statins increase the mRNA and the protein mass of HMGR, which translates to an over expression of cholesterol biosynthesis in intact lens (Cenedella 1995; Cenedella 1997). It is suggested that IP products might prevent lens opaqueness, cataract, and lens cholesterol deposition. Cataract removal remains the most common surgery in the US (more than half million per year). The occurrence of cataracts approaches 50% for those 75 years or older. The protective use of Vitamin E against cataract development is well recognized (VERIS 1990). Vitamin E tocotrienols and tocopherols are both powerful antioxidants. However, only tocotrienols, especially delta- and gamma-tocotrienols have been shown to down regulate the mRNA and reduce the protein mass of HMGR.